Reducing Supply Chain Dependencies for Viral Genomic Surveillance: Get by with a Little HELP from Commercial Enzymes already in your Lab Freezer

<ns3:p>Background The COVID-19 pandemic exposed critical vulnerabilities in global laboratory supply chains, disrupting the availability of key reagents and jeopardising the continuity of genomic surveillance essential for epidemic response. Sustaining sequencing capacity during supply shortages requires practical, locally accessible alternatives to commercial kits. Methods We developed ARTIC HELP (Homebrew Enzymes for Library Preparation), an open-source adaptation of the ARTIC nanopore sequencing protocol for viral genomic surveillance. We described cost-effective, generic replacements for all enzyme mixes used in tiling multiplex RT-PCR and the nanopore native barcoding workflow, including end-prep (EP), barcode ligation (BL), and adapter ligation (AL). Through systematic evaluation, we tested wild-type M-MLV reverse transcriptase and two types of proofreading DNA polymerases, (i) B-family Pfu-based polymerases fused to an Sso7d DNA-binding domain, and (ii) blends of A-family (Taq-based) and B-family (Pfu-based) polymerases, against standard reagents. We validated the workflow on clinical SARS-CoV-2 and Norovirus GII samples. Results The HELP workflow delivered genome coverage comparable to the ARTIC LoCost protocol. For SARS-CoV-2 samples (Ct ≤28), wild-type M-MLV RT combined with selected Pfu or AB blend polymerases, alongside optimized HELP EP, BL, and AL mixes, achieved 84.0–99.6% genome coverage. For Norovirus GII samples (Ct ≤32), the HELP workflow enabled >85% coverage across six of eight genotypes tested. While some polymerases showed reduced performance at higher Ct values, they performed reliably at Ct <24, supporting their use as emergency alternatives in rapid outbreak-response sequencing when viral load is high and RNA quality sufficient. Conclusions ARTIC HELP provides a robust and flexible solution to maintain viral sequencing capacity when standard reagents are inaccessible or unaffordable. Our cost analysis highlights stark global disparities in reagent pricing driven by import fees and supply barriers, rather than protocol complexity, underscoring the need for equitable pricing models and local sourcing strategies. By giving laboratories more options for essential reagents, ARTIC HELP strengthens preparedness and resilience for future public health emergencies.</ns3:p>